title: Use of anion-aromatic interactions to position the general base in the ketosteroid isomerase active site.
abstract: Although the cation-pi pair, formed between a side chain or substrate cation and the negative electrostatic potential of a pi system on the face of an aromatic ring, has been widely discussed and has been shown to be important in protein structure and protein-ligand interactions, there has been little discussion of the potential structural and functional importance in proteins of the related anion-aromatic pair (i.e., interaction of a negatively charged group with the positive electrostatic potential on the ring edge of an aromatic group). We posited, based on prior structural information, that anion-aromatic interactions between the anionic Asp general base and Phe54 and Phe116 might be used instead of a hydrogen-bond network to position the general base in the active site of ketosteroid isomerase from Comamonas testosteroni as there are no neighboring hydrogen-bonding groups. We have tested the role of the Phe residues using site-directed mutagenesis, double-mutant cycles, and high-resolution X-ray crystallography. These results indicate a catalytic role of these Phe residues. Extensive analysis of the Protein Data Bank provides strong support for a catalytic role of these and other Phe residues in providing anion-aromatic interactions that position anionic general bases within enzyme active sites. Our results further reveal a potential selective advantage of Phe in certain situations, relative to more traditional hydrogen-bonding groups, because it can simultaneously aid in the binding of hydrophobic substrates and positioning of a neighboring general base.
authors.affiliation: Department of Biochemistry, Stanford University, Stanford, CA 94305, USA.
authors.forename: Jason P
authors.initials: JP
authors.lastname: Schwans
authors.forename: Fanny
authors.initials: F
authors.lastname: Sunden
authors.forename: Jonathan K
authors.initials: JK
authors.lastname: Lassila
authors.forename: Ana
authors.initials: A
authors.lastname: Gonzalez
authors.forename: Yingssu
authors.initials: Y
authors.lastname: Tsai
authors.forename: Daniel
authors.initials: D
authors.lastname: Herschlag
mesh_terms: D000838:Anions; D020134:Catalytic Domain; D007664:Ketosteroids / Q000378:metabolism*; D009154:Mutation; D013251:Steroid Isomerases / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D014961:X-Ray Diffraction
chemical_list: D000838:Anions; D007664:Ketosteroids; D013251:Steroid Isomerases
keywords: enzyme catalysis; general-base catalysis; noncovalent interactions